Millions of people are infected with the AIDS virus HIV-1, and highly active antiviral therapy has made great strides over the past decade to slow virus spread and improve infected individuals' prognoses. Because many of the utilized anti-viral compounds elicit significant side effects, and rapid virus evolution leads to significant numbers of drug resistant strains, there is an ongoing need to discover new drugs to inhibit HIV-1 replication. Such research is driven through detailed understanding of the cellular and molecular biological steps that the virus undergoes to grow. The pivotal step in the viral replication cycle is the integration of the viral DNA made by reverse transcription into a cell chromosome. Integration is catalyzed by integrase, and the first integrase inhibitor was approved for clinical use in 2007. Herein, we will ascertain critical unknown aspects of integrase structure and function. Our work and others have previously highlighted that integrase works in close association with cellular factors to accomplish virus integration. The key factor, lens epithelium-derived growth factor (LEDGF), tethers HIV-1 to active genes during integration, but the underlying mechanistic basis of LEDGF tethering is unknown. This will be deciphered using a variety of cell biology, biochemical, and virological experimental techniques. Even in the complete absence of LEDGF, HIV-1 still favors active genes for integration over random, leading to the hypothesis that other virus-interacting proteins help to guide HIV-1 as it seeks chromosomal sites. The roles of numerous other viral binding proteins in integration will therefore be deciphered. Though it is believed that a tetramer of the integrase protein is the multimer that catalyzes integration, a dearth of detailed structural biology information has limited our overview of its organization. Biochemical and virological experiments will be conducted to define the functional organization of the active integrase tetramer. As LEDGF is the natural tether that guides the virus to sites of integration, novel DNA binding domains will be tested in concert with LEDGF to ascertain the extent that HIV-1 can be directed to new sites for integration. The successful completion of these experiments will uncover fundamental aspects of HIV- 1 integrase structure and function, which will significantly help in the discovery of novel integrase inhibitors. They moreover may open up new ways in which to develop lentiviral vectors for future treatment of patients with genetic therapy. PUBLIC HEALTH RELEVANCE: HIV-1 replication critically relies on integrase activity, the viral enzyme that integrates the reverse transcript into a cell chromosome. This application will uncover novel aspects of integrase structure and function, which will significantly impact ongoing discovery efforts to target and block the pivotal integration step in the virus replication cycle.